Initiation of anterior pattern formation in moth fly embryos by Opa/Zic

Unrelated transcription factors initiate anterior pattern formation in morphologically similar embryos of different fly species. Apart from Bicoid in the model organism Drosophila melanogaster, it is not known how these so-called anterior determinants (ADs) establish the embryo's head-to-tail polarity. We examined how odd-paired, the AD gene of the moth fly Clogmia albipunctata, affects chromatin accessibility and gene expression during the early phase of zygotic genome activation. To answer this question, we assembled and annotated the genome of Clogmia albipunctata, performed single embryo ATAC-seq, and identified putative cis-regulatory elements in the Clogmia embryo during the last four nuclear division cycles of the blastoderm stage. We then established a protocol for studying chromatin accessibility and gene expression changes in single blastoderm embryos that were subjected to RNA interference (RNAi), using Clogmia's ortholog of the pioneer factor gene zelda (Cal-zld) as target, which had a pronounced impact on chromatin accessibility and gene expression. Finally, we examined how knockdown of maternal odd-paired transcript (Cal-opaMat) affected chromatin accessibility and gene expression during early stages of zygotic genome activation and identified homeobrain and sloppy-paired orthologs as its earliest transcription factor gene targets.